Neuro2a (N2a) mouse neuroma cells

Cell Introduction

cloneNeuro-2Ais caused byR.J.KlebeandF.H.RuddlesutraAEstablishment of self generated tumors in white mice. This cell produces a large amount of microtubule protein, which plays a role in the contraction system of the axonal flow response in nerve cells.

Cell characteristics

1）Source:Neuroblastoma of the brain

2）form：Amoeba like stem cells

3）Content:>1x10⁶ Cell count

4）SpecificationsT25bottleor1mLFreezing tube packaging

5) Purpose:For scientific research purposes only.

Transportation and storage:dry icetransportationAnd revive well to survivecell：（1）1mLFrozen storage tube packaging for dry ice transportation, upon receipt-80After overnight storage in the refrigerator, transfer to liquid nitrogen or directly resuscitate,If dry ice is found to have evaporated completelyThe bottle cap of the cryovial falls offDamaged and contaminated cells, please contact us immediatelycontact（2）T25Survival of Bottle ResuscitationcellShipped at room temperature,harvestFollow the processing method after receiving the cells.

After cell receptionHandling of:

1）After receiving the cells,75%Alcohol disinfection bottle wall willT25Bottle placed37Place the incubator at approximately ℃2-3hIf you find any damage to the culture bottle, overflow of liquid, or contamination of cells, please take photos and contact us promptly.

2）Please in4or5XConfirm the cell status under the microscope and take photos of the newly received cells（10×，20(X) Each2-3Please keep a photo of Zhang and the appearance of the culture bottle as a reference for the cell status upon receipt during after-sales service.

3）Adherent cells: Cells in37Placed in a ℃ incubator2-3hObserving the growth and adhesion of cells under a microscope, some adherent cells may detach and form clusters due to vibration during express delivery. If the growth density of cells is observed under a microscope, then60%The following can remove the culture medium from the culture bottle (if there are non adherent cells, they need to be recovered by centrifugation and resuspended in the original culture bottle),Add newly prepared specialized culture medium6-8mLContinue to culture in the cell culture incubator. If the cell growth density reaches70%-80%Above, cells can be passaged. During the passage process, if cells shed due to transportation vibrations, they need to be recovered by centrifugation.

4）Note: The transport medium (infusion medium) cannot be used to culture cells anymore. Please use a specialized medium newly prepared according to the instructions for cell culture conditions to culture cells. Suggestions for the first passage after receiving cellsT25culture flask1：2subculture

Oneculture mediumandcultivatecryopreservationCondition preparation:

1）prepareMEMCulture medium;premiumFetal bovine serum,10%；Dual antibody,1%.

2）culture conditionsGas phase: air,95%Carbon dioxide,5%Temperature:37℃，The humidity of the incubator is70%-80%.

3）cryopreservation solution：90%serum，10%DMSO，Currently usednowmatch.

IIcellhandle：

1）Recovery of frozen cells：：

Will contain1mLThe freezing tube for cell suspension37Quickly shake and thaw in a ℃ water bath, then add to the solution containing4-6mL dedicatedMix the culture medium evenly in the centrifuge tube. in1000RPMCentrifuge under conditions3-5minDiscard the supernatant and resuspend the cells in specialized culture medium. Then add the cell suspension to the solution containing6-8ml dedicatedIn the culture bottle (or dish) of the culture medium37Incubate overnight at ℃. The next day, observe cell growth and cell density under a microscope.

2）Cell passage:ifcellDensity reaches80%-90%，namelycan be performedsubculture.

For the passage of adherent cells, the following methods can be referred to:

1. discardCultivate the supernatant and use it without calciumMagnesium ionofPBSMoisturizing cells1-2time.

2. join0.25％（w / v）Trypsin-0.53 mM EDTAYuCultivation bottle（T25bottle1-2mL，T75bottle2-3mL），placed in37℃incubatorMiddle digestion1-2minute(Difficult to digest cells can be appropriately prolonged for digestion time)And thenObserve under a microscope afterwardscell digestionsituation,ifMost cells become roundAnd fall offQuickly retrieve the console and tap lightlya few timesAfter culturing the bottlejoin3-4mlcontain10%FBSThe cultivation mediumterminatedigestion.

3.Gently mix and suck out, in1000RPMitemCentrifuge under the piece3-5minDiscard the supernatant and add more1-2mLBlow the culture medium evenly.Press the cell suspension1：2The proportion is allocated to the newT25Bottle, add6-8mlAccording to the instructions, configure a new specialized culture medium to maintain the growth vitality of cells. Subsequent passages will be conducted according to the actual situation1:2~1:5The proportion is carried out.

3）Cell cryopreservation:It is recommended to receive the cells before cultivation3Freeze a batch of cell seeds for future experiments. belowT25Taking bottles as an example;

1. cellWhen frozenaccording toThe process of cell passage involves collecting digested cells into centrifuge tubes, which can be counted using a hemocytometer to determine the freezing density of the cells. The recommended freezing density for general cells is1×10⁶~1×10⁷A living cell/ml.

2. 1000rpmcentrifugation3-5minRemove the supernatant. Resuspend the cells in the prepared cell cryopreservation solution, according to each step1mlFreezing solution contains1×10⁶~1×10⁷A living cell/mlAssign cells to a cryovial and label them with information such as name, generation, date, etc.

3. Place the cells to be frozen in a programmed cooling box,-80Store overnight in the refrigerator and then transfer to a liquid nitrogen container for storage. Simultaneously record the position of the cryovial in the liquid nitrogen container for future reference and use.

Notes:

1. All animal cells are considered to have potential biological hazards and must be operated in a secondary biosafety platform. Please pay attention to protection, and all waste liquids and containers that have come into contact with these cells must be sterilized before disposal.

2. It is recommended to always use protective gloves, clothing, and a face mask when reviving frozen cells. Attention: The cryotube immersed in liquid nitrogen will leak and gradually fill with liquid nitrogen. When thawing, the conversion of liquid nitrogen into gas phase may cause the container to explode or the lid to be blown off with dangerous force, resulting in flying debris and causing personal injury.